Hetero-oligomeric CPN60 resembles highly symmetric group-I chaperonin structure revealed by Cryo-EM.

The chloroplast chaperonin system is indispensable for the biogenesis of Rubisco, the key enzyme in photosynthesis. Using Chlamydomonas reinhardtii as a model system, we found that in vivo the chloroplast chaperonin consists of CPN60α, CPN60β1 and CPN60β2 and the co-chaperonin of the three subunits CPN20, CPN11 and CPN23. In Escherichia coli, CPN20 homo-oligomers and all possible other chloroplast co-chaperonin hetero-oligomers are functional, but only that consisting of CPN11/20/23-CPN60αβ1β2 can fully replace GroES/GroEL under stringent stress conditions. Endogenous CPN60 was purified and its stoichiometry was determined to be 6:2:6 for CPN60α:CPN60β1:CPN60β2. The cryo-EM structures of endogenous CPN60αβ1β2/ADP and CPN60αβ1β2/co-chaperonin/ADP were solved at resolutions of 4.06 and 3.82 Å, respectively. In both hetero-oligomeric complexes the chaperonin subunits within each ring are highly symmetric. Through hetero-oligomerization, the chloroplast co-chaperonin CPN11/20/23 forms seven GroES-like domains, which symmetrically interact with CPN60αβ1β2. Our structure also reveals an uneven distribution of roof-forming domains in the dome-shaped CPN11/20/23 co-chaperonin and potentially diversified surface properties in the folding cavity of the CPN60αβ1β2 chaperonin that might enable the chloroplast chaperonin system to assist in the folding of specific substrates.